A major limitation to effective targeted gene repair is the low efficiency of homology-directed repair in most human cell types. The mechanism of homology-directed repair is now understood at a very sophisticated level of detail, and the goal of this component of the Northwest Genome Engineering Consortium (NGEC) is to harness this mechanistic understanding to develop methods for stimulating HDR at target genes. We describe three specific aims to achieve this goal: (1) We will develop a convenient reporter assay for homology-directed repair, and use it to optimize repair at double-strand breaks, in cell culture models and in primary hematopoietic cells. (2) We will ask if targeted gene repair can be efficiently initiated at nicks; and develop a reporter assay for break-induced genomic instability to establish whether nicks provide a safety advantage over double-strand breaks by diminishing the potential for translocation in therapeutic applications. (3) We will establish how chromatin structure contributes to gene marking and to targeted gene repair, and ask specifically if chromatin structure can be manipulated to increase the efficiency of gene repair. These planned experiments complement and mesh with other components of the NGEC to achieve the long-term goal of establishing methods for effective gene therapy of monogenic disorders of hematopoietic cells.